Membrane and cytoskeletal changes associated with IgE-mediated serotonin release from rat basophilic leukemia cells

Binding of antigen to IgE-receptor complexes on the surface of RBL-2H3 rat basophilic leukemia cells is the first event leading to the release of cellular serotonin, histamine, and other mediators of allergic, asthmatic, and inflammatory responses. We have used dinitrophenol-conjugated bovine serum albumin (DNP-BSA) as well as the fluorescent antigen, DNP-B-phycoerythrin, and the electron-dense antigen, DNP-BSA-gold, to investigate dynamic membrane and cytoskeletal events associated with the release of [3H]serotonin from anti-DNP-IgE-primed RBL-2H3 cells. These multivalent antigens bind rapidly to cell surface IgE-receptor complexes. Their distribution is initially uniform, but within 2 min DNP-BSA-gold is found in coated pits and is subsequently internalized. Antigen internalization occurs in the presence and absence of extracellular Ca2+. The F-actin content of the detergent-extracted cell matrices analyzed by SDS PAGE decreases during the first 10-30 s of antigen binding and then increases by 1 min to almost double the control levels. A rapid and sustained increase is also observed when total F-actin is quantified by flow cytometry after binding of rhodamine-phalloidin. The antigen-stimulated increase in F-actin coincides with (and may cause) the transformation of the cell surface from a finely microvillous to a highly folded or plicated topography. Other early membrane responses include increased cell spreading and a 2-3-fold increase in the uptake of fluorescein-dextran by fluid pinocytosis. The surface and F-actin changes show the same dependence on DNP-protein concentration as stimulated [3H]serotonin release; and both the membrane responses and the release of mediators are terminated by the addition of the non-cross-linking monovalent ligand, DNP-lysine. These data indicate that the same antigen-stimulated transduction pathway controls both the membrane/cytoskeletal and secretory events. However, the membrane and actin responses to IgE-receptor cross-linking are independent of extracellular Ca2+ and are mimicked by phorbol myristate acetate, whereas ligand-dependent mediator release depends on extracellular Ca2+ and is mimicked by the Ca2+ ionophore A23187.     

Subcellular Distribution and Developmental Expression of Cholesterol Ester Hydrolases in Fetal Rat Brain Cultures

Abstract: Cholesterol ester hydrolase activities previously have been identified in brain and linked to the production of myelin, which has very low levels of esterified cholesterol. We have studied two cholesterol ester hydrolase activities (termed the pH 6.0 and pH 7.2 activities) in cultures derived from 19- to 21-day-old dissociated fetal rat brains and in developing rat brain. In vivo the levels of both the pH 6.0 and pH 7.2 activities began to increase by about 10 postnatal days, reached maximal levels at 20 days (20 and 1.5 nmol/h/mg protein, respectively), and thereafter remained nearly constant (pH 6.0) or decreased somewhat before becoming constant (pH 7.2). In contrast, in the cultures the pH 6.0 cholesterol ester hydrolase activity was low until 21 days in culture (DIC; 20 nmol/h/mg protein), increased to a peak activity at 31 DIC (60 nmol/h/mg protein), remained high for 24 days, and finally decreased (18 nmol/h/mg protein at 63 DIC); the pH 7.2 cholesterol ester hydrolase activity was very low until 20 DIC, increased to a peak activity at 31 days (3 nmol/h/mg protein), and thereafter decreased to a lower level (2 nmol/h/mg protein) that was maintained for about 24 days before decreasing (0.7 nmol/h/mg protein at 63 DIC). Therefore, (a) the time courses of appearance of both cholesterol ester hydrolase activities were delayed by 10–14 days relative to that seen in vivo, and (b) the specific activities observed in the cultures were transiently two- to three-fold higher than in rat brain, but then declined to levels characteristic of whole brain homogenates. Subcellular fractionation of the cultures demonstrated that the pH 7.2 cholesterol ester hydrolase activity, along with myelin basic protein and 2′,3′-cyclic nucleotide-3′-phosphohydrolase activity, was enriched in a membrane fraction collected at an interface between 0.32 M and 0.9 M sucrose; the pH 6.0 cholesterol ester hydrolase activity, in contrast, was enriched in the microsomal fraction.     

Increased sensitivity to somatostatin in obese Zucker rats

The release of somatostatin from the pancreas and stomach following the ingestion of a meal and its increase in the peripheral circulation elicits an attenuation of postprandial hormone secretion such as insulin, pancreatic polypeptide and gastrin and retards the rate at which nutrients enter the circulation. Reduced tissue somatostatin content and/or an attenuated somatostatin release is associated with hyperinsulinism and obesity in certain animal models. In the obese Zucker rat, however, tissue somatostatin levels are increased and therefore the present study was designed to determine the effect of synthetic somatostatin on basal and postprandial arterial insulin levels in obese and lean Zucker rats. Synthetic somatostatin was infused at doses of 0.25, 0.5, 1 and 5 ng/kg X min before and after the intragastric instillation of a liver extract/sucrose test meal. In the obese rats somatostatin at a dose of 5 ng/kg X min reduced basal plasma insulin levels significantly, whereas no effect of somatostatin was observed on basal insulin levels in the lean animals at all doses employed. The integrated postprandial insulin response was reduced during 0.25, 0.5, 1 and 5 ng/kg X min somatostatin in the obese animals, whereas only 0.5 ng/kg X min and higher doses had an inhibitory effect in the lean rats. The degree of inhibition in relation to the postprandial insulin response during saline infusions was 35-230% in the obese and 30-100% in the lean Zucker rats within the range of somatostatin infusions employed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Progenitors of oligodendrocytes: Limiting dilution analysis in fetal rat brain culture

In this paper, we describe the use of a combination of cell culture techniques and limiting dilution analysis to determine the number of oligodendrocyte progenitor cells and the oligodendrocyte clone size in primary dispersed cultures of 20- to 21-day-old fetal rat brain. Single-cell suspensions (1,2,3 × 10⁶ cells/ml) were plated in either microwell or 100 mm dishes. After 22 days in culture the number of differentiated oligodendrocytes was ascertained by determining the amount of myelin basic protein by radioimmunoassay. The total amount of myelin basic protein was the same in the two types of dish, indicating that proliferation and differentiation were unaffected when oligodendrocytes were grown in microwells. The fraction (F₀) of microwells containing no oligodendrocytes was determined at each cell dilution. F₀ decreased exponentially with increasing total cell concentration. The linearity of the plot of ln F₀ versus cell number indicates that the number of oligodendrocyte progenitor cells is limiting. From the equation describing the Poisson distribution of progenitor cells in microwells we calculated that, at the time of plating, primary cultures of fetal rat brain contain one oligodendrocyte progenitor cell per 1.3 × 10⁵ brain cells, or a total population of 300–500 progenitor cells per brain. The mean oligodendrocyte clone size was determined to be approximately 825 at 22 days and close to 2000 by 35 days in culture. Therefore, each progenitor cell must undergo approximately 11 divisions, on the average, during postnatal development.     

Das geruchsorgan der rezenten Actinistia und Dipnoi (Pisces)

The olfactory organs of all recent genera from the Actinistia (Latimeria) and Dipnoi (Neoceratodus, Lepidosiren, Protopterus) were studied morphologically, and compared with each other. Whereas the olfactory organ of Latimeria resembles that of the Brachiopterygii, the olfactory organ of the Dipnoi is similar to that, of the Elaemohra.nnhii and ArtinnnfarAraii

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Nit dankenswerter Unterstützung durch die Deutsche Forschungsgemeinschaft.     

Deletion der kurzen Arme des Chromosoms Nr. 18

Ohne ZusammenfassungDirektor: Prof. Dr. Dr. W. Lenz